Lyme disease is one of most prevalent vector-borne human diseases in the United States, Europe and Asia. According to the United States Centers for Disease Control, the rate of Lyme disease infection in the United States is about 7.9 cases for every 100,000 persons. In some areas, such as New England, the infection rate reaches as high as 31.6 cases for every 100,000 persons. The disease is transmitted by the Ixodes ricinus ticks, which carry the spirochete Borrelia species including Borrelia burgdorferi or Borrelia afzelii. Through the tick bites, the spirochetes eventually affect multiple organs of the body. Lyme disease is clinically manifested by dermatologic (e.g., erythema migrans), cardiac, neurologic (e.g., meningitis), rheumatologic and musculoskeletal (e.g., joint pain) abnormalities. Left untreated, infected patients may acquire chronic lyme borreliosis. It is estimated that 61% of patients with erythema migrans would eventually develop neurologic, articular or cardiac symptoms.
In spite of the great demand, early detection for Lyme disease remains elusive. There are only few commercial tests currently available. In addition, significant limitations exist in these diagnostic methods, which include bacterial cultures, serologic methods, and genetic detection via polymerase chain reaction (PCR). See, A. C. Steere, N. Engl. J. Med. 321, 568-596 (1989). For example, culturing of Borrelia species from clinical samples is proven to be time consuming and tedious. Serologic detection of Lyme disease by Western Blot analysis is unreliable. Cross-reactions with other spirochetes are seen in some tests. In Western Blot analysis, purified microbial proteins are separated by SDS-PAGE and followed by detection using Lyme-specific anti-sera. However, antigenic variations among patients can make the test unreliable. Appearance of IgG and IgM titers in patients after infection may fluctuate greatly (e.g., IgG peaks 3-12 months and IgM peaks 3-6 weeks). Different titer responses among patients may yield false negative results, drastically limiting the accuracy of the serologic method.
U.S. Pat. No. 6,087,097 discloses detection of B. burgdorferi using a PCR assay based on specific genomic/plasmid DNA sequences of OspA gene from B. burgdorferi. However, this PCR-based assay has gained only limited success, in part because of its lack of specificity. Because of this limitation, the PCR technique has not become a standardized routine diagnosis.
Because ticks (e.g., I. persulcatus) may carry multiple spirochete Borrelia species, including B. afzelii and B. garinii, either singly or simultaneously, a combination of microorganisms may simultaneously invade a human body through a single tick bite. There are three established sub-groups of Borrelia bacteria: (i) Borrelia garinii (European “sub-group”) which causes neurological illnesses; (ii) Borrelia afzelli (European “sub-group”) which causes Acrodermatis atrophicans chronic and (iii) Borrelia burgdorferi (American “sub-group”) which causes especially Arthritis and rheumatic pains).